The aim of this research is to select out and characterize early and late stages in the evolution of neoplasia in the respiratory epithelium. Our unique approach is to expose the respiratory epithelium to the carcinogen, DMBA, in vivo using the rat tracheal implant model which permits carefully controlled exposure to a well delineated target. Specific cell populations will be identified in a nondestructive manner by cutting the tracheal implant into explants, placing them into short-term organ culture, and identifying the lesions on the explants by the cells which exfoliate into the medium. The lesions will be classified according to severity based on criteria commonly used in human cytopathology. Our first set of experiments is designed to test whether an increase in severity of the lesion (mild-moderate-marked atypias, etc.) as determined cytopathologically, correlates with a decrease in time to the acquisition of in vivo markers of neoplasia (survival under selective stringent culture conditions, anchorage-independent growth, etc.) and tumorigenicity during cell culture of epithelial cell populations, derived from specific lesions. These studies will validate the experimental approach and provide biological evidence for the sequential cytopathological classification of the lesions which presently is inferred from human biopsy and autopsy specimens. The second study is to determine whether a known tumor promoter, TPA, can enhance the progression of neoplasia in cell populations derived from cytologically undetectable as well as mild to marked atypias, i.e. from any stage in the development of cancer. This study will provide a system for testing other agents that may enhance and, more important to preventive medicine, inhibit particular stages in the development of neoplasia. Our third set of experiments is aimed at determining the biochemical basis for the early changes in growth requirements (loss of requirement for insulin and pyruvate) seen in carcinogen preexposed tracheal cells. Changes in uptake of hexoses into cell populations derived from tumors, selected lesions, and controls will be measured, as well as the influence of insulin on this uptake. These will provide biochemical markers of stages in the development of lung cancer and form a baseline for studying the molecular alterations in the cells (plasma membrane?) in the future.